الفهرس 
chapter I: Introduction And Aim Of The WorkOnly 14 pages are availabe for public view
 |  | from | 127 |  |  |
| | | from | 127 | | |
Abstract
Toxoplasma gondii (T. gondii), an obligate intracellular parasite, has the unusual ability of being able to invade the nucleated cells of warm-blooded animals. Over one billion people are estimated to be infected with this parasite worldwide. Human infection by T. gondii is generally asymptomatic and induces a self-limiting disease in immunocompetent individuals. However, the effects of infection are much more severe in immunocompromised individuals. In addition, primary T. gondii infection acquired during pregnancy can be transmitted to the fetus and can cause severe diseases like miscarriage, neurological damage, and ocular complications. Additionally, the consumption of food contaminated with tissue cysts of T. gondii, such as meat from infected livestock, is the main route of parasite transmission to humans.
For toxoplasmosis treatment, the recommended drugs have side effects and reactivation may occur at any time. Nevertheless, there are not any therapies, which can eradicate this organism from the host. On the other hand, immunization against T. gondii is another way to control this infection. Therefore, the development of a safe and effective vaccine would be extremely valuable in fighting against T. gondii infection.
In recent years, significant progress has been made in the identification of vaccine candidates against toxoplasmosis that could elicit a protective immune response. At present, the only accepted vaccine against toxoplasmosis is ”Toxovax” which contains a live attenuated S48 strain that controls congenital infection in ewes. Toxovax decreases the abortion rate but does not eradicate the parasite. However, it is expensive and may be changed into a pathogenic form therefore, it is not appropriate for human use. Furthermore, there is no licensed vaccine for humans, Therefore, the development of an effective and safe vaccine against T. gondii would be of great value for the control of the parasite infection in humans and animals.
The objective of this study was to identify and characterize a 44-KDa target antigen extracted from the T. gondii tachyzoite and investigate its vaccine potential.
Separation of the target antigen from the rapidly growing phase (Tachyzoite) using electrophoresis technique on polyacrylamide gel (SDS-PAGE) and dyeing it using Coomassie blue. Also, determination of the molecular weight of the target antigen using the western blot technique by using a highly Specific Rabbit IgG polycolonal antibody to Toxoplasma Ags to identify the target antigen at 44-KDa.
The target antigen was also purified using a polyacrylamide elution gel, and a partial molecular biochemical characterization of the antigen was done during its purification.
BALB/C mice (n=20) were randomly divided into two groups, each group containing 10 mice. the group I mice were injected with the target antigen, the group II were injected with PBS . Blood samples were collected from mice, separated, and the serum was obtained.
Survival days are the most important parameters to consider to assess the protective effect of native 44-KDa vaccines against the T. gondii challenge ,so, Four groups of BALB/c mice were challenged with a lethal dose of 6.45 x 103 tachyzoites of T. gondii RH strain to evaluate the survival rate , each group (n=10) . First group immunized with PBS , Second group immunized with 44-KDa toxoplasma antigen , groups III immunized with 44-KDa antigen then intraperitoneally challenged with a lethal dose of 6.45 x103 live T.gondi RH strain tachyzoites, and group IV were intraperitoneally challenged with a lethal dose of 6.45 x103 live T.gondi RH strain tachyzoites . All mice were observed for 21 days after challenge at 3-day intervals in each group.This experiment was repeated once to verify the results .
Using a particular antibody, Target TAg44 was partially identified as a protein after purification.
High levels of IgG antibodies were detected in the sera of immunized mice with the 44-kDa antigen.
In the sera of mice immunized with the 44-kDa Toxoplasma antigen, high amounts of both IgG1 and IgG2a were discovered, indicating that both Th1 and Th2 type responses were induced.
Significant levels (p>0.001) of interferon-gamma INF-γ (Th1) and significant levels (p<0.001) of interleukin 10 (IL-10) (Th2) being produced in serum samples in mice immunized with 44-KDa. These finding confirming the results of the IgG subclass that 44-kDa purified Toxoplasma antigen can induce both arms of immune response (Th1 & Th2).
In the present study, BALB/c mice were challenged with a lethal dose of 6.45 x 103 tachyzoites of T. gondii RH strain to evaluate the survival rate.
All mice of infected (challenged) group died within 6-12 days after the RH tachyzoite challenge. The group mice immunized with 44-KDa showed longer survival time after being infected with RH tachyzoites that achieved (70%) survival rates. Therefore, the 44-KDa produced considerable protection, which could partially prevent mice from T. gondii fatal infection.